Experiments will be carried out to determine whether the altered properties of I-cell disease (ICD) and pseudo-Hurler polydystrophy (PHP) lysosomal hydrolases involve an alteration of a common post-translational pathway (possibly glycosylation) specific for the lysosmal hydrolases. Included among these studies are the purification and characterization of alpha-L-fucosidase, N-acetyl-beta-D-hexosaminidase and alpha-D-glucosidase from normal ICD and PHP urine samples; carbohydrate analysis and thin layer peptide mapping of these purified proteins and interaction of these glycoproteins with Concanavalin A. The purified normal, ICD and PHP enzymes will be used to investigate the specific uptake and/or lack of such uptake by cultured human fibroblasts, Kupffer cells, hepatocytes, aveolar macrophages and the interaction of the enzymes with the mannan specific binding protein from rabbit liver. Elucidation of the structural requirements for the specific recognition and uptake of lysosomal glycohydrolases will be carried out using glycopeptides prepared from purified N-acetyl Beta-D-hexosaminidase and alpha-L-glucosidase. Tunicamycin will be used to ascertain whether an alteration in carbohydrate is responsible for the mislocalization of several lysosomal enzymes to the extracellular culture medium. Glycopeptide profiles from the purified ICD and PHP enzymes will be compared to the corresponding normal samples. Genetic and biochemical complemenation studies should resolve the biochemical heterogeneity that is characteristic of these disorders while cell fusion experiments may allow us to map the gene responsible for the molecular defect in ICD and PHP.